The long-term objectives of the proposed research are to define the intracellular signaling mechanisms through which contractile and growth stimuli elicit their effects in vascular smooth muscle (VSM). A detailed understanding of these pathways could provide a basis for explaining the pathology underlying pervasive vascular diseases such as hypertension, atherosclerosis, and injury-induced myointimal hyperplasia. This knowledge could also provide sites for the rational targeting of therapeutic agents aimed at controlling VSM cell proliferation and contractile function. Protein kinase C (PKC) is an enzyme which is an important component of intracellular signaling pathways in diverse tissues and cells. Work funded in years 1-5 of this grant, has implicated PKC as a factor in the regulation of VSM contractile activity. We have determined that multiple PKC isozymes are expressed in VSM, providing a basis for stimulus-specific patterns of PKC activation and for explaining diverse PKC actions. In this application: I) We propose to test the hypothesis that the activities of specific PKC isozymes in cultured VSM are differentially regulated by stimulation of intact cells with phorbol ester activators of PKC and physiological growth and contractile stimuli. 2) We intend to utilize this information to determine the role of specific PKC isozymes in mediating the activation of a signaling pathway, involving mitogen-activated protein kinase (MAP-kinase), by contractile and growth stimuli in these cells. Activation of this signaling pathway is required for proliferative cell growth. 3) Recent data suggest that a signaling pathway involving MAP-kinase may also be operative in fully differentiated VSM and serve a role in contractile regulation. If so, information gained in the cultured cell system, pertaining to the role of PKC isozymes in mediating MAP kinase activation and cell growth responses, may be applicable and important for a full understanding of contractile regulation in intact arterial tissue. Therefore, the role of MAP- kinase in mediating phorbol ester-stimulated contractile responses will be determined in intact arterial smooth muscle and the contribution of PKC to the activation of MAP-kinase in intact tissues by contractile stimuli will be determined. 4) Finally, we will test the hypothesis that cGMP-dependent and cAMP-dependent vasodilators will inhibit MAP kinase activation, contributing to subsequent relaxation of contractile activity in intact tissue and inhibition of cell growth in the cultured cells. If proven true, the specific step in the signaling pathway which is inhibited will be determined.